The long-term goal of this application is to elucidate the fundamental mechanism by which dysregulation of the histone demethylase, GASC1 (Gene Amplified in Squamous Cell Carcinoma 1, also known as JMJD2C and KDM4C), contributes to tumorigenesis, and to lay a foundation for the development of this protein as a new therapeutic target against breast cancer. The GASC1 gene was originally cloned from an amplified region at 9p24 in esophageal cancer cells. Later studies demonstrated that GASC1 is amplified in approximately 15% of breast cancers with overexpression more prevalent in aggressive, basal-type breast cancer. The GASC1 protein is a key member of histone demethylases that play an essential role in regulating chromatin architecture and gene expression; and is implicated in tumorigenesis. GASC1 mainly catalyzes demethylation of tri- and di-methylated forms of histone H3 lysine 9 (H3K9me3/me2) epigenetic repressive marks. However, the molecular mechanisms by which GASC1-dependent chromatin regulation translates to oncogenicity and cancer progression remain poorly understood. Intriguing preliminary evidence indicated that GASC1 is significantly enriched at target gene promoter regions, and that recruitment of GASC1 to specific genomic loci requires the GASC1 Tudor and Plant Homeo Domains (PHD). Studies indicate that the Tudor and PHD domains have the potential to bind H3K4me3 active marks at promoter regions. Importantly, we demonstrated that GASC1 target genes are involved in multiple signaling pathways and biological processes, including critical genes such as the S-phase kinase-associated protein 2 (SKP2) which participates in ubiquitination. The central hypothesis of this application is that GASC1 is recruited to gene promoter regions containing H3K4me3 active marks via its histone-binding domains, and the subsequent demethylation of H3K9me3/me2 repressive marks induces the transcription of a set of key ubiquitination pathway genes that ultimately promote tumorigenesis. Based on this hypothesis, strategies that selectively alter the GASC1 histone recruitment hold great promise as targeted therapies for aggressive, GASC1-amplified breast and esophageal cancers. In this application, two specific aims will be pursued. In Aim 1, we will elucidate the molecular mechanism and structural details of GASC1 that mediate its recruitment to promoter regions of target genes. In Aim 2, we will determine how GASC1 impacts the histone methylation status and expression of target genes responsible for mediating GASC1's role in breast tumorigenesis in vitro and in xenograft animal models. Significantly, the proposed research will fundamentally increase our understanding of the mechanisms by which GASC1 is recruited to genomic loci and how genetic amplification of GASC1 alters epigenetic programming and triggers downstream oncogenic pathways in cancer. These aspects have translational implications in the development of GASC1 mechanism-based therapies to target a wide range of cancers, particularly GASC1-amplified basal breast cancer.